| Objective:To study the optimum technology for purification of total lignans from Herpetospermum caudigerum,and the protective effects of Herpetospermum polysaccharide on immunologic liver injury induced by concanavalin A ?ConA? in mice was investigated simultaneously.Methods: ? The isolation and purification of ethyl acetate extract from Herpetospermum caudigerum were conducted by means of ordinary pressure normal phase silica gel column chromatography,Sephadex LH-20,preparative HPLC etc., The structures of the compounds were identified based on their physical and chemical features,and spectral data.? The method on content determination for total lignans in Herpetospermum caudigerum was established by UV-VIS spectrophotometry, while that of herpetone was established by HPLC-DAD.? The statically saturated adsorption capacity and desorption capacity were compared with seven different types of macroporous resin and polyamide resin with the content of herpetone and total lignans as the targets,so as to screen out the appropriate resin category. Then,investigations of singal fator was used to select optimum technological parameters to ascertain the best purification process for the total ligans.? In this part,Herpetospermum polysaccharide was used as the drug on the treatment of immunological liver injury in mouse which were injected ConA through tail vein. And then, liver index and spleen index,contents of alanine aminotransferase?ALT?, aspartate aminotransferase?AST?, latic dehydrogenase ?LDH?, nitrogen monoxidum?NO? and inter leukin-6?IL-6? in serum, superoxide dismutase?SOD?, malondialdehyde?MDA? in liver homogenate and pathological morphological changes in liver cells were monitored.Results and Conclusion: ? Seven compounds were isolated from this extractive of them were elucidated as herpetolide B??? n herpetolide A???? balanophonin????herpetone????cassyformin ???,compounds ??? and ? were lignans,and compounds ? and ? were isolated from this plant for the first time.? The content of total lignans was determined by the chromotropic acid-H2SO4 reaction colorimetry,measurement wavelength was set at 566 nm,and the amount of H2SO4 was 7 mL,the amount of chromotropic acid was 1.5 mL,and reaction time was 40 minutes. The total lignans and herpetone calibration curves showed a good liner relationship in ranger of 5.010-17.535 ?g·mL-1?n=6,r=0.9991?,0.0501-0.5010 ?n=6,r=0.9999? ?g·mL-1 respectively. The methods established were reliable and accurate.They can be used to determine the content of herpetone and total lignans in Herpetospermum caudigerum.? The HP-20 macroporous resin was selected to separate and purify lignans compounds from Herpetospermum caudigerum,and the best technology conditions as follow:the sample concentration was 0.8 g?native herbs?·mL-1 with a folllow rate at 1 BV·h-1,the ratio of diameter to height of resin column was 1:8,the sample volume was 0.8 g·mL-1 ?native herbs/wet resin?, the amount of water was 3 BV, the total lignans were eluted by 80% alcohol with a folllow rate at 2 BV·h-1, and the amount of 80% alcohol was 6 BV. After purification, the purity of total lignans and herpetone were 79.18%,0.97%, separately,increased significantly than before. This study could provide a reference for the production of total lignans from Herpetospermum caudigerum.? Herpetospermum polysaccharide has a good effct on immunological liver injury in mouse,the mechanism possibly due to its function of anti-inflammatory,anti-oxidation,scavenging oxygen free radical and regulating immunity. |
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